Variations in Membrane Sensitivity of Brain Region Synaptosomes to the Effects of Ethanol In Vitro and Chronic In Vivo Treatment

The effects of chronic ethanol treatment on the membrane order of synaptosomes from the cerebral cortex, striatum, cerebellum, brainstem, and hippocampus of rats were determined by measuring the fluorescence polarization of diphenylhexatriene (DPH) that had been incorporated into the synaptosomal membranes. Fischer-344 rats either were fed a nutritionally complete ethanol-containing liquid diet for 5 months or pair-fed with a diet that contained sucrose substituted isocalorically for ethanol. Polarization values for synaptosomes from all the brain regions studied were similar except for those from cerebral cortical synaptosomal membranes, which were significantly less ordered. Ethanol in vitro (30-500 mM) decreased the polarization values in synaptosomes from sucrose-control rats for all brain regions, although the sensitivity of cerebellar synaptosomes to the membrane disordering effects of ethanol in vitro was significantly greater that of synaptosomes from other brain regions. Chronic ethanol treatment did not alter baseline polarization for any brain region. Cerebellar and brainstem synaptosomes from the ethanol-fed rats were significantly less susceptible to the membrane disordering effects of ethanol in vitro compared to their sucrose controls, suggesting that chronic ethanol administration results in tolerance to ethanol's membrane effects. Striatal synaptosomes exhibited intermediate tolerance, whereas the sensitivities of cortical and hippocampal synaptosomes to membrane disordering by ethanol in vitro were not significantly affected by the chronic ethanol treatment. These results suggest that synaptosomal membranes have different membrane order requirements depending on the brain region from which they are prepared. Variations in brain regional neuronal membrane sensitivity to ethanol and differential tolerance development may contribute to some of the acute and chronic behavioral effects of ethanol.     

Differential Regulation of Phosphoinositide Phosphodiesterase Activity in Brain Membranes by Guanine Nucleotides and Calcium

We have shown previously that calcium and guanine nucleotides stimulate the activity of a phosphoinositide (PI) phosphodiesterase in membranes from rat cerebral cortex and that their effects are additive. To understand further guanine nucleotide- and calcium-stimulated PI phosphodiesterase activity, we have investigated the pH sensitivity and effects of inhibitors on the two modes of stimulation. NaF stimulates PI hydrolysis in brain membranes with an EC50 of 2 mM and a maximal effect at 10 mM, suggesting that a guanine nucleotide binding protein can regulate PI phosphodiesterase. Neomycin inhibited guanylylimidodiphosphate (GppNHp)-stimulated PI phosphodiesterase activity in a concentration-dependent manner, with 90% inhibition at 0.3 mM. Neomycin was not as effective at inhibiting calcium-dependent PI hydrolysis (32% inhibition at 0.3 mM). Chloroquine also had a greater inhibitory effect against GppNHp-stimulated PI phosphodiesterase activity compared to calcium-dependent activity. Guanine nucleotide- and NaF-dependent activations of PI phosphodiesterase were strongly pH-dependent, with greatest stimulation observed at pH 5-6 and inhibition at more alkaline pH. Calcium-stimulated PI hydrolysis was not as sensitive to changes in pH and had a peak of activity at pH 9. Our findings of different pH optima and differential sensitivity to inhibitors suggest that calcium and guanine nucleotides may regulate PI phosphodiesterase in rat cortical membranes through independent mechanisms.     

Cholinergic- and Adrenergic-Stimulated Inositide Hydrolysis in Brain: Interaction, Regional Distribution, and Coupling Mechanisms

Carbachol and norepinephrine were used as agonists to compare and contrast cholinergic and adrenergic stimulation of inositide breakdown in rat brain slices. Carbachol acts through a muscarinic (possibly M1) receptor and norepinephrine acts through an alpha 1 adrenoceptor. Studies in cerebral cortical slices indicated that both agonists stimulated the production of inositol-1-phosphate and glycerophosphoinositol. Although the initial rates for the stimulation of inositol phosphate release were similar for the two ligands, the response to norepinephrine continued for 60 min and was larger compared with carbachol which plateaued at 30 min. The presence of carbachol did not affect the ED50 for norepinephrine. Concentrations of carbachol near the ED50 in combination with norepinephrine resulted in an additive response whereas maximal concentrations of carbachol and norepinephrine resulted in a less than additive response in the cortex. This negative interaction was also seen in the hippocampus and hypothalamus but not in the striatum, brainstem, spinal cord, olfactory bulb, or cerebellum. Norepinephrine had a larger response than carbachol in the hippocampus, striatum, and spinal cord, but the reverse was true in the olfactory bulb. Manganese (1 mM) stimulated the incorporation of [3H]inositol into phosphatidylinositol (PtdIns) four- to fivefold but not into polyphosphoinositides. The stimulation by manganese of PtdIns labelling increased the nonstimulated release of inositol phosphates but did not affect the stimulated release of inositol phosphates by carbachol or norepinephrine.(ABSTRACT TRUNCATED AT 250 WORDS)     

Seasonal fluctuations of resource abundance and avian feeding guilds across forest–farmland boundaries in tropical Africa

Seasonal fluctuations in climatic factors are expected to increase in future decades. However, little is known about the response of tropical species communities to seasonal fluctuations in climate and resource availability, particularly across different habitat types. We examined the relationship between spatio‐temporal fluctuations in the abundance of fruits and invertebrates and two avian feeding guilds, i.e. frugivores and insectivores, in forest and farmland habitats in western Kenya. Fruits and invertebrates fluctuated substantially throughout the year, but seasonal fluctuations were asynchronous between the two habitat types. Species richness and total abundance of frugivores and insectivores also fluctuated strongly and were closely related to the abundance of their respective resources. Frugivore species richness fluctuated anti‐cyclical in forest and farmland habitats, suggesting that several frugivorous species tracked fruit resources across habitat boundaries. In contrast, insectivorous bird richness fluctuated synchronously in the two habitat types, suggesting a lack of local‐scale movements across habitat boundaries. We conclude that bird communities strongly respond to seasonal fluctuations in resource availability, but responses differ between feeding guilds. While frugivores seem to respond flexibly to seasonal fluctuations, for instance by tracking fruit resources across habitat boundaries, insectivorous birds appear to be more susceptible to the expected increase in seasonal fluctuations in resource availability.     

Radical disinfestation protocol eliminates in vitro contamination in Guava (<Emphasis Type="Italic">Psidium guajava </Emphasis>L.) seeds

Controlling contamination in vitro is one of the basic requirements for successful tissue culture technology. In preliminary studies, in vitro contamination of guava seeds was almost 100%, even when disinfested with bleach. To overcome this problem, we developed a unique method to control contamination using bleach and strong acids. Submerging guava seeds in 10% HCl for 24–72 h followed by a 30 min treatment with 10% bleach (NaOCl) reduced contamination rates from 98 to 0%. Substituting 5% H₂SO₄ for 12 h for 10% HCL gave similar results. In addition to eliminating contamination, acid-treated seeds germinated faster in vitro than control seeds germinated in a greenhouse (15 vs. 40 days, respectively).     

High Bird Species Diversity in Structurally Heterogeneous Farmland in Western Kenya

Tropical ecosystems are globally important for bird diversity. In many tropical regions, land‐use intensification has caused conversion of natural forests into human‐modified habitats, such as secondary forests and heterogeneous agricultural landscapes. Despite previous research, the distribution of bird communities in these forest‐farmland mosaics is not well understood. To achieve a comprehensive understanding of bird diversity and community turnover in a human‐modified Kenyan landscape, we recorded bird communities at 20 sites covering the complete habitat gradient from forest (near natural forest, secondary forest) to farmland (subsistence farmland, sugarcane plantation) using point counts and distance sampling. Bird density and species richness were on average higher in farmland than in forest habitats. Within forest and farmland, bird density and species richness increased with vegetation structural diversity, i.e., were higher in near natural than in secondary forest and in subsistence farmland than in sugarcane plantations. Bird communities in forest and farmland habitats were very distinct and very few forest specialists occurred in farmland habitats. Moreover, insectivorous bird species declined in farmland habitats whereas carnivores and herbivores increased. Our study confirms that tropical farmlands can hardly accommodate forest specialist species. Contrary to most previous studies, our findings show that structurally rich tropical farmlands hold a surprisingly rich and distinct bird community that is threatened by conversion of subsistence farmland into sugarcane plantations. We conclude that conservation strategies in the tropics must go beyond rain forest protection and should integrate structurally heterogeneous agroecosystems into conservation plans that aim at maintaining the diverse bird communities of tropical forest‐farmland mosaics.     

Microdialysis of Ethanol During Operant Ethanol Self-administration and Ethanol Determination by Gas Chromatography

Operant self-administration methods are commonly used to study the behavioral and pharmacological effects of many drugs of abuse, including ethanol. However, ethanol is typically self-administered orally, rather than intravenously like many other drugs of abuse. The pharmacokinetics of orally administered drugs are more complex than intravenously administered drugs. Because understanding the relationship between the pharmacological and behavioral effects of ethanol requires knowledge of the time course of ethanol reaching the brain during and after drinking, we use in vivo microdialysis and gas chromatography with flame ionization detection to monitor brain dialysate ethanol concentrations over time. Combined microdialysis-behavioral experiments involve the use of several techniques. In this article, stereotaxic surgery, behavioral training and microdialysis, which can be adapted to test a multitude of self-administration and neurochemical centered hypotheses, are included only to illustrate how they relate to the subsequent phases of sample collection and dialysate ethanol analysis. Dialysate ethanol concentration analysis via gas chromatography with flame-ionization detection, which is specific to ethanol studies, is described in detail. Data produced by these methods reveal the pattern of ethanol reaching the brain during the self-administration procedure, and when paired with neurochemical analysis of the same dialysate samples, allows conclusions to be made regarding the pharmacological and behavioral effects of ethanol.     

Ethanol seeking by long evans rats is not always a goal-directed behavior

Background

Two parallel and interacting processes are said to underlie animal behavior, whereby learning and performance of a behavior is at first via conscious and deliberate (goal-directed) processes, but after initial acquisition, the behavior can become automatic and stimulus-elicited (habitual). With respect to instrumental behaviors, animal learning studies suggest that the duration of training and the action-outcome contingency are two factors involved in the emergence of habitual seeking of “natural” reinforcers (e.g., sweet solutions, food or sucrose pellets). To rigorously test whether behaviors reinforced by abused substances such as ethanol, in particular, similarly become habitual was the primary aim of this study.

Methodology/Principal Findings

Male Long Evans rats underwent extended or limited operant lever press training with 10% sucrose/10% ethanol (10S10E) reinforcement (variable interval (VI) or (VR) ratio schedule of reinforcement), or with 10% sucrose (10S) reinforcement (VI schedule only). Once training and pretesting were complete, the impact of outcome devaluation on operant behavior was evaluated after lithium chloride injections were paired with the reinforcer, or unpaired 24 hours later. After limited, but not extended instrumental training, lever pressing by groups trained under VR with 10S10E and under VI with 10S was sensitive to outcome devaluation. In contrast, responding by both the extended and limited training 10S10E VI groups was not sensitive to ethanol devaluation during the test for habitual behavior.

Conclusions/Significance

Operant behavior by rats trained to self-administer an ethanol-sucrose solution showed variable sensitivity to a change in the value of ethanol, with relative insensitivity developing sooner in animals that received time-variable ethanol reinforcement during training sessions. One important implication, with respect to substance abuse in humans, is that initial learning about the relationship between instrumental actions and the opportunity to consume ethanol-containing drinks can influence the time course for the development or expression of habitual ethanol seeking behavior.     

Population Genetics of Two Key Mosquito Vectors of Rift Valley Fever Virus Reveals New Insights into the Changing Disease Outbreak Patterns in Kenya

Rift Valley fever (RVF) outbreaks in Kenya have increased in frequency and range to include northeastern Kenya where viruses are increasingly being isolated from known (Aedes mcintoshi) and newly-associated (Ae. ochraceus) vectors. The factors contributing to these changing outbreak patterns are unclear and the population genetic structure of key vectors and/or specific virus-vector associations, in particular, are under-studied. By conducting mitochondrial and nuclear DNA analyses on >220 Kenyan specimens of Ae. mcintoshi and Ae. ochraceus, we uncovered high levels of vector complexity which may partly explain the disease outbreak pattern. Results indicate that Ae. mcintoshi consists of a species complex with one of the member species being unique to the newly-established RVF outbreak-prone northeastern region of Kenya, whereas Ae. ochraceus is a homogeneous population that appears to be undergoing expansion. Characterization of specimens from a RVF-prone site in Senegal, where Ae. ochraceus is a primary vector, revealed direct genetic links between the two Ae. ochraceus populations from both countries. Our data strongly suggest that unlike Ae. mcintoshi, Ae. ochraceus appears to be a relatively recent, single ''introduction'' into Kenya. These results, together with increasing isolations from this vector, indicate that Ae. ochraceus will likely be of greater epidemiological importance in future RVF outbreaks in Kenya. Furthermore, the overall vector complexity calls into question the feasibility of mosquito population control approaches reliant on genetic modification.     

Nature conservation at the edge

Currently, there is an increasing need for evidence-based strategies in nature conservation, for example when designing and establishing nature reserves. In this contribution, we critically assess the ecological relevance of recent nature conservation practices in Kenya (East Africa), a region of global biodiversity hotspots. More specifically, we overlay the distribution of species richness (here based on mammals, birds, amphibians and vascular plants) with the location of nature reserves, the Kenyan agro-ecological zones (areas representing diverging agricultural potentials), and with the spatial distribution of human population density. Our analyses indicate that the majority of protected areas are located in areas with comparatively low species richness, while areas with extraordinary high levels of species richness are not adequately covered by nature reserves. Areas of high agricultural productivity (and with high human demographic pressure) are mainly reserved for high-yield agriculture; however, these regions are also characterised by high species richness. The majority of nature reserves are restricted to the semi-arid regions of Kenya, marginal for agricultural usage, but also with low levels of species richness. Based on this analysis, we prioritize areas for future protection. This single-country case illustrates that agricultural production in high-yield areas outweighs nature conservation goals, even in global biodiversity hotspot regions, and that priority setting may conflict with effective nature conservation.